Abstract
Emphasis was placed in this work on the assessment of the role of guanine bases in the interaction of transcription factor Sp1 with its cognate DNA sequence. For this purpose, each guanine residue of the 5′-GGGGCGGGG-3′ (GC box) target DNA sequence was substituted in turn by 8-oxo-7,8-dihydro-2′-deoxyguanosine. The latter oxidized nucleotide which is likely to be present in mammalian DNA and exhibit mutogenic features is expected to be involved in age-related diseases and cancer. The effect of the incorporation of 8-oxodGuo into DNA on the binding of transcription factor Sp1 was studied using electrophoretic mobility shift assays with nuclear extracts from HeLa cells. When guanines at position G′2, G′3, G′5 and G′6 were replaced with 8-oxodGuo, binding of Sp1 was only 28%, 30%, 7%, 5% and 21%, respectively, to that of the non-substituted oligonucleotide. The binding is less affected when guanines at position G′1, G′7, G′8 and G′9 were substituted by 8-oxodGuo. Results show up the importance of the core of the GC box and the stronger contribution of the second and the third zinc finger to the binding with DNA. All together, this suggests that incorporation of 8-oxodGuo may alter the expression of the gene regulated by Sp1 and affect the response of the cell.